...Title of Experiment 9: Measurement of Nucleic Acid Solutions Objectives: 1. To study the principle of gel electrophoresis. 2. To use gel electrophoresis method to measure the nucleic acid solutions. 3. To learn the technique of nuclei acid measurement by using gel electrophoresis. Introduction: Gel electrophoresis is a widely used technique for the analysis of nucleic acids and proteins. Most every molecular biology research laboratory routinely uses agarose gel electrophoresis for the preparation and analysis of DNA. The components of gel electrophoresis system is power supply and a chamber, agarose gel which is a porous material that allows molecules migrate through, buffer with a mixture of water and ions, and gel casting tray and comb. Electrophoresis is a method of separating substances based on the rate of movement while under the influence of an electric field. Agarose is a polysaccharide purified from seaweed. An agarose gel is created by suspending dry agarose in a buffer solution, boiling until the solution becomes clear, and then pouring it into a casting tray and allowing it to cool. The result is a flexible gelatin-like slab. Nucleic acids are composed of chains of nucleotides. The ‘backbone’ of the nucleic acid structure is a repeating chain of phosphate groups and pentose sugars. At certain pH values, the oxygen atoms in the phosphate groups ionize, giving the molecule an overall negative charge. If exposed to an electric field, these molecules are attracted...
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...Agarose Gel Electrophoresis for the Separation of DNA Fragments Pei Yun Lee, John Costumbrado, Chih-Yuan Hsu, Yong Hoon Kim Department of Molecular, Cell, and Developmental Biology, University of California Los Angeles Video Article Chapters 0:05 Title 1:31 Preparation of the Gel 3:21 Setting up Gel Apparatus and Separating DNA Fragments 4:55 Observing Separated DNA Fragments 5:43 Results: Agarose Gel Electrophoresis of PCR Products 6:23 Conclusion Cite this Article Lee, P. Y., Costumbrado, J., Hsu, C. Y., Kim, Y. H. Agarose Gel Electrophoresis for the Separation of DNA Fragments. J. Vis. Exp. (62), e3923, doi:10.3791/3923 (2012). Abstract Agarose gel electrophoresis is the most effective way of separating DNA fragments of varying sizes ranging from 100 bp to 25 kb1. Agarose is isolated from the seaweed genera Gelidium and Gracilaria, and consists of repeated agarobiose (L- and D-galactose) subunits2. During gelation, agarose polymers associate non-covalently and form a network of bundles whose pore sizes determine a gel's molecular sieving properties. The use of agarose gel electrophoresis revolutionized the separation of DNA. Prior to the adoption of agarose gels, DNA was primarily separated using sucrose density gradient centrifugation, which only provided an approximation of size. To separate DNA using agarose gel electrophoresis, the DNA is loaded into pre-cast wells in the gel and a current applied. The phosphate backbone of the DNA (and RNA) molecule...
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...Green chemistry, also called sustainable chemistry, is a philosophy of chemical research and engineering that encourages the design of products and processes that minimize the use and generation of hazardous substances. Whereas environmental chemistry is the chemistry of the natural environment and of pollutant chemicals in nature, green chemistry seeks to reduce the negative impact of chemistry on environment by preventing pollution at its source and using fewer natural resources. As a chemical philosophy, green chemistry applies to organic chemistry, inorganic chemistry, biochemistry, analytical chemistry, physical chemistry and even chemical engineering. While green chemistry seems to focus on industrial applications, it does apply to any chemistry choice. Click chemistry is cited as a style of chemical synthesis that is consistent with the goals of green chemistry. The focus is on minimizing the hazard and maximizing the efficiency of any chemical choice. In 2005 three key developments in green chemistry were identified: use of superficial carbon dioxide as green solvent, aqueous hydrogen peroxide for clean oxidations and the use of hydrogen in asymmetric synthesis. Examples of applied green chemistry are supercritical water oxidation, on water reactions and dry media reactions. Bioengineering is also seen as a promising technique for achieving green chemistry goals. A number of important process chemicals can be synthesized in engineered organisms, such as shikimate, a Tamiflu...
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